POS0611 SHARED MONOCYTE CYTOKINE SIGNATURE INDUCED BY SERUM FROM PATIENTS WITH SYSTEMIC LUPUS ERYTHEMATOSUS AND ANTI-MDA5 ANTIBODY-POSITIVE DERMATOMYOSITIS THROUGH TYPE I INTERFERON PATHWAY
Authors: S. Nakamura, Y. Okamoto, Y. Katsumata, M. Harigai
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Type I interferon (IFN) plays a pivotal role in the pathogenesis of systemic lupus erythematosus (SLE) and dermatomyositis (DM)/polymyositis (PM) as typically evidenced by upregulation of type I IFN-stimulated genes (ISGs) in peripheral immune cells [1]. In addition, a distinct monocyte cytokine signature, which was induced by plasma from pediatric SLE patients and abrogated by Janus kinase (JAK) inhibition, was reported [2]. However, contribution of serum factors to expression of ISGs was not fully elucidated. <h3>Objectives</h3> To investigate the cytokine induction profile in multiple myeloid lineages by serum from patients with SLE or DM/PM using the whole blood stimulation system and identify the signaling pathway relavant to the monocyte cytokine signature. <h3>Methods</h3> Serum was collected from newly diagnosed, untreated, and clinically active adult patients with SLE (SLE group), anti-MDA5 antibody-positive DM (MDA5 group), and anti-aminoacyl-tRNA synthetase (ARS) antibody-positive DM/PM (ARS group), and healthy controls (HCs) (<i>n</i> = 10 for each group). Heparinized whole blood from healthy donors was incubated with control serum (10 v/v%), patient serum (10 v/v%), or IFN-β in the presence of protein transport inhibitor cocktail for 6 hours. Red blood cells were lysed, and then leucocytes were fixed at a single step. Intracellular staining was performed and expression of 11 cytokines in CD14<sup>+</sup> monocytes, CD1c<sup>+</sup> dendritic cells (DCs), and CD123<sup>+</sup> DCs were analyzed using flow cytometry. A cut-off level was determined by 2% positivity of each cytokine in unstimulated condition. For transcriptomic analyses, sorted CD14<sup>+</sup> monocytes from healthy donors were incubated with serum (SLE, MDA5, ARS, or HCs), or IFN-β (<i>n</i> = 3 for each group) for 4 hours, and bulk RNA-sequencing was performed using the Illumina NextSeq 500 platform. RNA-seq raw sequence reads analysis and pathway analysis were performed using the Strand NGS software. To evaluate significance of JAK-signal transducer and activator of transcription (STAT) pathway, whole blood from healthy donors was pre-incubated with upadacitinib, a JAK1 inhibitor, stimulated with patient serum for 6 hours, and analyzed for cytokine expression as described above. <h3>Results</h3> Serum from SLE and MDA5 groups induced significantly higher monocyte chemoattractant protein-1 (MCP1) and interleukin-1 receptor antagonist (IL-1RA) expression in CD14<sup>+</sup>monocytes than serum from HCs (Figure 1). These monocyte cytokine signatures were closely resembled that induced by IFN-β stimulation. Serum from ARS group did not induce any significant cytokine expression in CD14<sup>+</sup> monocytes. No significant cytokine expression was observed in CD1c<sup>+</sup> DCs or CD123<sup>+</sup> DCs. RNA-seq demonstrated that 612 and 462 genes were upregulated (fold change &gt;2 relative to control) following stimulation with serum from SLE and MDA5 groups, respectively. In these upregulated genes, 383 genes were commonly upregulated across SLE and MDA5 groups and IFN-β. Pathway analysis revealed similar transcriptional profiles in SLE and MDA5 groups: upregulated genes were most frequently involved in IFN-αβ signaling pathway including multiple ISGs and STAT1/2. Upadacitinib significantly abrogated the monocyte cytokine signature, and MCP1 and IL-1RA expression induced by serum from SLE and MDA5 groups in a dose-dependent manner (<i>p</i> &lt; 0.001 in all analyses). <h3>Conclusion</h3> These results suggest that serum factors in patients with active SLE and anti-MDA5 antibody-positive DM can induce shared monocyte cytokine signature through type I IFN pathway. CD14<sup>+</sup> monocytes 'primed' by serum might contribute to the pathogenesis of these diseases. <h3>References</h3> [1]<i>Rheumatology (Oxford).</i> 2017;56:1662 [2]<i>J Autoimmun</i>. 2017;81:74 <h3>Acknowledgements:</h3> NIL. <h3>Disclosure of Interests</h3> Shohei Nakamura: None declared, Yuko Okamoto: None declared, Yasuhiro Katsumata Speakers bureau: GlaxoSmithKline K.K., AstraZeneca K.K., Sanofi K.K., Pfizer Japan Inc., Janssen Pharmaceutical K.K., Chugai Pharmaceutical Co., Ltd., Asahi Kasei Pharma, Astellas Pharma Inc., Mitsubishi Tanabe Pharma Corporation, Masayoshi Harigai: None declared.